This invention relates to methods and compositions for treating and/or preventing Pneumocystis carinii infections in mammals.
P. carinii pneumonitis (PCP) is one of the leading causes of death of victims of acquired immunodeficiency syndrome (AIDS). Untreated, the mortality rate of PCP in AIDS patients approaches 100%. During 1986, the number of deaths from PCP in the United States exceeded the combined number of deaths from all types of meningococcal infections, viral hepatitis and encephalitis, gonorrhea, syphilis, varicella, measles, mumps, rubella, diphtheria, tetanus, pertussis, polio, amebiasis, shigellosis, salmonellosis, typhoid fever, typhus fever, cholera, rabies, brucellosis, anthrax, tularemia, botulism, and malaria. The clinical course of PCP infection in immunosuppressed patients presents a number of symptoms including tachypnea, cough, fever, hypoxemia (blood oxygen deficiency), an increased alveolar-capillary oxygen gradient, respiratory acidosis and bilateral diffuse alveolar disease (Hughes, W. T., N. Eng. J. Med., 317: 1021-1023, 1987).
PCP is caused by a ubiquitous lung dwelling organism. Recent evidence indicates that this pathogen may be a fungus rather than a protozoan as previously thought (Meshnick, S. et al., "Antioxidant Enzymes of Pneumocystis carinii", Abstracts from the 36th Annual Meeting of the American Society of Tropical Medicine and Hygiene, p. 235, 1987; Edman, J. C. et al., Nature 334: 519-522, 1988; Stringer et al., "Sequence from Ribosomal RNA from Pneumocystis carinii compared to those of Four Fungi Suggests an Ascomycetous Affinity", Journal of Protozoology 36: 14S-16S, 1989; Watanabe et al., "5S Ribosomal RNA Sequence of Pneumocystis carinii and its Phylogenetic Association with )Rhizopoda/Myxomycota/Zygomycota Group", Journal of Protozoology 36: 16S-18S, 1989; Edman et al., "Ribosomal RNA Genes of Pneumocystis carinii", Journal of Protozoology 36: 18S-20S, 1989).
Early potential treatments for the disease were tested on the infantile P. carinii interstitial plasma-cell pneumonitis that occurred in epidemic form in Europe. The mortality rate for the untreated form of the affliction was about 50%. At that time, it was determined that various antibiotics and/or anti-microbial agents, such as penicillin, tetracycline, chloramphenicol, streptomycin, quinacrine, chloroquine, neoarsphenamine, stibophen, pamaquine, mulsin, neospiran, arsaphen, arsphenamine, quinine, chloroguanide and emetine hydrochloride were ineffective in treating the disease (Hughes, W., Pneumocystis carinii Pneumonitis, CRC Press Incorporated, 1987).
Defined high risk groups exist for PCP. While the ubiquitous P. carinii acts as a commensal organism and does not cause disease in a healthy individual, it may nevertheless produce PCP in, for example, patients immunosuppressed due to AIDS, to drugs given for cancer treatment, to drugs given to prevent rejection of organ or tissue transplant, to drugs given for treatment of autoimmune disease and other immunocompromised, i.e., partially or totally immunosuppressed or immunodeficient, patients or hosts. Because PCP poses such a threat to these identifiable high risk groups, there exists a need for a prophylactic routine as well as a therapeutic treatment.
The development of treatments for PCP has been hindered by the lack of knowledge about the biology and properties of the P. carinii organism. The nutritional requirements, metabolic pathways, mode of replication, enzyme systems and taxonomy of P. carinii are not well understood. Although there are some newly identified agents active against PCP, only two recognized treatments currently exist for the control of PCP.
The first recognized mode of treatment involves the use of pentamidine, p,p'-(pentamethylenedioxy)dibenzamidine bis(beta-hydroxy-ethanesulfonate) which is an aromatic diamidino compound. Pentamidine may be prepared according to the disclosure in Newberry Easson, U.S. Pat. No. 2,410,796 and is available commercially, e.g., from LyphoMed, Inc., Rosemont, Ill. 60018).
The second treatment involves the use of a combination of trimethoprim with sulfamethoxazole (TMP/SMZ), i.e., 5-[(3,4,5-trimethoxyphenyl)methyl]-2,4-pyrimidinediamine/4-amino-N-(5-meth yl-3-isoxazolyl)benzenesulfonamide). TMP may be prepared from guanidine and beta-ethoxy-3,4,5-trimethoxybenzylbenzalnitrile (see, e.g., Stenbuck, Hood, U.S. Pat. No. 3,049,544 and Hoffer, U.S. Pat. No. 3,341,541). SMZ may be prepared starting with ethyl 5-methylisoxazole-3-carbamate (see Kano et al., U.S. Pat. No. 2,888,455). TMP and SMZ, including the combination of TMP/SMZ, are available commercially from a number of sources. For example, a suspension of TMP/SMZ (400 mg/200 mg) is available from Geneva Generics, Inc., Broomfield, Colo. 80020; in tablet form, TMP/SMZ may be obtained from Par Pharmaceutical, Inc., Spring Valley, N.Y. 10977. Other forms of TMP/SMZ-including commercial sources may be found by referring to Physician's Desk Reference, 1988 Edition, Med. Econ. Co., Inc., Oradell, N.J., p.325, col. 2.
The efficacy of the combination of TMP/SMZ derives from the ability of TMP to inhibit microbial dihydrofolate reductase activity and from the competitive interference of SMZ with the incorporation of para-aminobenzoic acid into dihydrofolate, which serves to limit the formation of substrate for the enzyme. Known disadvantages of these treatments include lack of clinical responsiveness, high rates of toxicity and numerous other adverse side-effects. In AIDS patients, in particular, the severe adverse reactions caused by pentamidine therapy include neutropenia, thrombocytopenia, rash and alterations in mental state, e.g., depression. Symptoms resulting from pentamidine therapy include hypoglycemia, hypotension and nephrotoxicity. In addition, it has been determined that AIDS patients suffering from PCP require therapy far longer periods of time and have higher relapse rates (Havertos, H. W., Am. J. Med. 76: 501-508, 1984).
New treatments undergoing clinical trials include various agents with modes of action similar to that of TMP/SMZ; i.e., interference with folate metabolism. These include: TMP in combination with dapsone, diaminodiphenylsulfone, a drug used to treat leprosy (Green et al., "AIDS-Related Pneumocystis carinii Pneumonia Successfully Treated with Dapsone-Trimethoprim", British Journal of Clinical Pharmacology 26: 487-491, 1988); trimetrexate (a new anti-cancer drug) in combination with leucovorin as a rescue agent for host metabolism (Allegra et al., "Trimetrexate for the Treatment of Pneumocystis carinii Pneumonia in Patients with Acquired Immunodeficiency Syndrome", New England Journal of Medicine 317: 978-985, 1987); high doses of steroids combined with specific anti-PCP therapy (Gallacher et al., "Treatment of Acute Pneumocystis carinii Pneumonia with Corticosteroids in a Patient with Acquired Immunodeficiency Syndrome", CritiCal Care Medicine 17: 104-105, 1989); administration of TMP/SMZ with careful monitoring of the serum concentration in individual patients during treatment so as to minimize adverse side effects (Sattler et al., "Trimethoprim-Sulfamethoxazole Compared with Pentamidine for Treatment of Pneumocystis carinii Pneumonia in the Acquired Immunodeficiency Syndrome", Annals of Internal Medicine 109: 280-287, 1988). TMP/SMZ has been administered prophylactically to AIDS patients to prevent PCP similar to the protocol found to be successful for children undergoing treatment for leukemia. Pentamidine has been formulated as an aerosol for delivery directly to the lungs primarily as a prophylactic protocol (Kovacs and Masur, "Pneumocystis carinii Pneumonia: Therapy and Prophylaxis", Journal of Infectious Diseases 158: 254-259, 1988). DL-alphadifluoromethylornithine (DFMO, eflornithine) is based on an entirely new mode of action and is under clinical evaluation for treatment of PCP (Schechter et al., "Clinical Aspects of Inhibition of Ornithine Decarboxylase with Emphasis on Therapeutic Trials of Eflornithine (DFMO) in Cancer and Protozoan diseases", in Inhibition of Polyamine Metabolism eds: McCann, Pegg and Sjoerdsma, Academic Press, pages 345-364, 1987). In addition to the new treatments described above, several others have been found to be active in animal models of PCP. These include: compounds related to pentamidine such as berenil (Clarkson et al., "Efficacy of DL-alphadifluoromethylornithine in a Rat Model of Pneumocystis carinii Pneumonia", Antimicrobial Agents and Chemotherapy 32: 1158-1163, 1988) and 1,4-di(4'-amidinophenoxy)butane (Tidwell et al., "Treatment of Experimental Pneumocystis carinii Pneumonia with Analogues of Pentamidine", Journal of Protozoology 36: 74s-77s, 1989); piritrexim, another anti-cancer drug which interferes with folate metabolism and is closely related to trimetrexate (Queener et al., "Activity of Lipid-Soluble Inhibitors of Dihydrofolate. Reductase against Pneumocystis carinii in Culture and in a Rat Model of Infection", Antimicrobial Agents and Chemotherapy 31: 1323-1327, 1987); and a combination of clindamycin and primaquine (Queener et al., "Activity of Clindamycin and Primaquine against Pneumocystis carinii in vitro and in vivo", Antimicrobial Agents and Chemotherapy 32: 807-813, 1988.
It has long been known that in many instances hosts and pathogens compete for trace nutrients such as iron, and that the ability of a pathogen to establish infection is often dependent on the ability of the pathogen to compete successfully against the host for trace nutrients (Jones, R. and Grady, R. W., Eur. J. Clin. Microbiol. 2:411-413, 1983). In order to secure needed iron, many microorganisms produce and release extremely effective, low molecular weight iron chelators known as siderophores, which compete with host iron-binding proteins. The microbe-produced siderophores allow the pathogens to survive the hypoferemia the host produces in response to infection . The host increases the amount of iron-binding proteins such as lactoferrin and transferrin which sequester iron making it unavailable to the microbe. When the microbe-produced siderophores complex with iron, the complex can be taken up by the microbe thus satisfying its nutritional requirement for iron.
Accordingly, it is known that a low bioavailable iron level may contribute to reducing susceptibility to certain microorganisms. It has also been reported that administration of iron compounds to patients suffering from certain diseases aggravates the condition (Masawe, A. E. J., et al., Lancet, 2: 314-317, 1974). Therefore, iron chelators, which act to complex iron, have long been known to have therapeutic potential. One use of iron chelators has been to treat iron overload in patients receiving multiple blood transfusions (such as those suffering from beta-thalassemia). Iron chelators have also been used as antibacterial and antimicrobial agents. For example, hydroxamic acids, a specific class of iron chelator, have been found to inhibit malaria sporozoites (Hynes, J. B., J. Med. Chem. 13: 1235-1237, 1970). Iron chelators have also been used in conjunction with antimicrobial agents in the control of Stapbylococcus epidermidis, Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis, and species of Salmonella, Enterobacter, Pseudomonas and Providencia (van Asbeck, B. et al., Eur. J. Clin. Microbiol. 2: 432-438, 1983). Although iron chelators have been employed to treat various microbial infections, their use is by no means predictable or uniformly effective. As an example, desferrioxamine, a hydroxamic acid iron chelator, has been shown to increase the virulence of S. typhimurium in mice (Jones, R. and Grady, R. W., supra).
Desferrioxamine (DFO) is known to be useful in the treatment of other illness; its ability to chelate aluminum has been exploited in the treatment of Alzheimer's disease (Mclachlan, U.S. Pat. No. 4,419,365 issued Dec. 6, 1985). It has also been used to suppress Plasmodium falciparum malaria (Pollack, S. et al., Proc. Soc. Exp. Diol. Med. 184: 162-164, 1987).